Higf 1 quantikine elisa

Reagents were obtained from Sigma (Munich, Germany) unless otherwise indicated. Recombinant FGF-2 and TGFβ3 were purchased from Peprotech (Hamburg, Germany). The dimethylmethylene blue dye was from Serva (Heidelberg, Germany). The anti-IGF-I (AF-291-NA) antibody was from R&D Systems (Wiesbaden-Nordenstadt, Germany), the anti-type-II collagen (II-II6B3) antibody from the NIH Hybridoma Bank (University of Iowa, Ames, IO, USA), the anti-type-I (AF-5610) antibody from Acris (Hiddenhausen, Germany), the anti-bromodeoxyuridine (BrdU; BU-33) and anti-type X collagen (COL-10) antibodies from Sigma, and the anti-SOX9 (C-20), anti-CD34 (C-18), anti-CD71 (C-20), and anti-CD105 (T-20) antibodies from Santa Cruz Biotechnology (Heidelberg, Germany). Biotinylated secondary antibodies and ABC reagent were obtained from Vector Laboratories (Alexis Deutschland GmbH, Grünberg, Germany). The IGF-I enzyme-linked immunosorbent assay (ELISA) (hIGF-I Quantikine ELISA) was from R&D Systems, the type II and type I collagen ELISAs (Arthrogen-CIA Capture ELISA Kit) from Chondrex (Redmond, WA, USA), and the type X collagen ELISA from Antibodies-online GmbH (Aachen, Germany). The Cell Proliferation reagent WST-1 was from Roche Applied Science (Mannheim, Germany) and the alkaline phosphatase (ALP) staining kit was from Sigma.

Recombinant TGF‐β (rTGF‐β) was purchased at Peprotech (Hamburg, Germany), dimethylmethyleneblue (DMMB) at Serva (Heidelberg, Germany) and diaminobenzidine (DAB) at Sigma‐Aldrich (Munich, Germany). The anti‐IGF‐I (AF‐291‐NA) antibody was from R&D Systems (Wiesbaden, Germany), the anti‐type‐II collagen (II‐II6B3) antibody from NIH Hybridoma Bank (University of Iowa, Ames, USA), the anti‐SOX9 (C‐20) antibody from Santa Cruz Biotechnology (Heidelberg, Germany), the anti‐type‐I collagen (AF‐5610) antibody from Acris (Hiddenhausen, Germany) and the anti‐type‐X collagen (COL‐10) antibody from Sigma‐Aldrich. Secondary biotinylated antibodies and the ABC kit were from Vector Laboratories (Alexis Deutschland GmbH, Grünberg, Germany). The hIGF‐I Quantikine ELISA was purchased at R&D Systems and the type‐II, ‐I and ‐X collagen ELISAs at Antibodies‐Online (Aachen, Germany).